There are a number of diseases of the central nervous system (“CNS”) which have a devastating effect on patients. These diseases are incurable and debilitating. They include Alzheimer's disease, Huntington's disease, Parkinson's disease and Multiple Sclerosis, to name a few.
By way of example, Parkinson's disease is a progressive degenerative disorder of unknown cause. In healthy brain tissue, dopaminergic neurons extend from the substantia nigra of the brain into the striatum. Parkinson's disease occurs when these dopaminergic neurons die. There are a number of methods to treat Parkinson's disease.
One method is to treat humans having parkinsonism with L-DOPA. Another method is to transplant cells into the substantia nigra or striatum. Transplanted cells replace endogenous cells that are lost as a consequence of damage. Transplanted cells may also be used as vectors for the expression of therapeutic molecules. Another method is to implant fetal brain grafts containing dopaminergic neurons. This method is experimental (Widner et al., 1993; Callahan et al., 1992). An animal model of Parkinson's disease is an MPTP-treated non-human primate. The animal models have been transplanted with dopamine-rich embryonic neurons with some success (Dunnett et al., 1991). (MPTP is a selective dopaminergic toxicant that produces parkinsonian symptoms in humans and in primates after a one-hit lesion to the neurons in the substantia nigra (Langston et al., 1983; Burns et al., 1983)).
Investigators studying other neurodegenerative diseases, such as Alzheimer's disease and Huntington's disease, are exploring the possible usefulness of fetal-tissue implants in the treatment of these diseases.
Current approaches to transplantation suffer from a number of serious limitations. First, many investigators are utilizing non-neural cells such as fibroblasts or transformed cell lines for transplantation. Second, the safety of transplantation of immortalized cell sources into the human brain is a concern. These cells may become unregulated and develop into tumors. Third, transplants of dopaminergic neuron fetal tissue to Parkinson's disease patients have a number of difficulties:                the fate of implanted dopaminergic neurons in patients with Parkinson's disease is uncertain—whatever caused the loss of endogenous dopaminergic neurons may also eventually injure the implanted ones,        in many cases, implants provide only temporary relief as the symptoms associated with the disease often return after a number of years,        the patient may reject foreign fetal tissue,        there are adverse reactions associated with immunosuppression (immunosuppression is needed to try to help the patient accept the foreign fetal tissue, even though the brain is, to some degree, immunologically privileged),        a sufficient number of cells in the fetal tissue being implanted are unable to survive during and after implantation,        the implants may not be regulated by the host brain,        other diseases or disorders may be transmitted to the patient via the implant,        the cost and effort associated with implanting fetal tissue may not be justified by the results, and        there are objections to the ethics associated with implanting fetal tissue.        
Many of these problems are encountered with transplants used to treat other neurodegenerative diseases, disorders or abnormal physical states.
In some tissues, stem cells and progenitor cells are proposed as a source for alternative treatments of disease or injury to tissues. The proposed treatments involve transplants of healthy tissue or endogenous stimulation of stem cells or progenitor cells to produce healthy tissue.
Stem cells are undifferentiated cells that exist in many tissues of embryos and adult mammals. In embryos, blastocyst stem cells are the source of cells which differentiate to form the specialised tissues and organs of the developing fetus. In adults, specialised stem cells in individual tissues are the source of new cells which replace cells lost through cell death due to natural attrition, disease or injury. No stem cell is common to all tissues in adults. Rather, the term “stem cell” in adults describes different groups of cells in different tissues and organs with common characteristics.
Stem cells are capable of producing either new stem cells or cells called progenitor cells. A progenitor cell differentiates to produce the mature specialized cells of mammalian organs. In contrast, stem cells never terminally differentiate (i.e. they never differentiate into specialized tissue cells). Progenitor cells and stem cells are referred to collectively as “precursor cells”. This term is often used when it is unclear whether a researcher is dealing with stem cells or progenitor cells or a combination of both cells.
Progenitor cells may differentiate in a manner which is unipotential or multipotential. A unipotential progenitor cell is one which can form only one particular type of cell when it is terminally differentiated. A multipotential progenitor cell has the potential to differentiate to form more than one type of tissue cell. Which type of cell it ultimately becomes depends on conditions in the local environment such as the presence or absence of particular peptide growth factors, cell—cell communication, amino acids and steroids. For example, it has been determined that the hematopoietic stem cells of the bone marrow produce all of the mature lymphocytes and erythrocytes present in fetuses and adult mammals. There are several well-studied progenitor cells produced by these stem cells, including three unipotential and one multipotential tissue cell. The multipotential progenitor cell may divide to form one of several types of differentiated cells depending on circumstances such as which hormones or factors act upon it and cell—cell contact.
Weiss et al, 1996, summarises the five defining characteristics of stem cells as the ability to:                Proliferate: Stem cells are capable of dividing to produce daughter cells.        Exhibit self-maintenance or renewal over the lifetime of the organism: Stem cells are capable of reproducing by dividing symmetrically or asymmetrically to produce new stem cells. Symmetric division occurs where one stem cell divides into two daughter stem cells. Asymmetric division occurs where one stem cell forms one new stem cell and one progenitor cell. Symmetric division is a source of renewal of stem cells. This permits stem cells to maintain a consistent level of stem cells in an embryo or adult mammal.        Generate large number of progeny: Stem cells may produce a large number of progeny through the transient amplification of a population of progenitor cells.        Retain their multilineage potential over time: Stem cells are the ultimate source of differentiated tissue cells, so they retain their ability to produce multiple types of progenitor cells, which will in turn develop into specialized tissue cells.        Generate new cells in response to injury or disease: This is essential in tissues which have a high turnover rate or which are more likely to be subject to injury or disease, such as the epithelium or blood cells.        
Thus, the key features of stem cells are that they are multipotential cells which are capable of long-term self-renewal over the lifetime of a mammal.
There has been much effort to isolate stem cells and determine which peptide growth factors, hormones and other metabolites influence stem cell renewal and production of progenitor cells, which conditions control and influence the differentiation of progenitor cells into specialized tissue cells, and which conditions cause a multipotential progenitor cell to develop into a particular type of cell.
Stem cells or progenitor cells may be used as substrates for producing healthy tissue where a disease, disorder or abnormal physical state has destroyed or damaged normal tissue. For example, stem cells and progenitor cells may be used as a target for in vivo stimulation with growth factors or they may be used as a source of cells for transplantation. The stem cells or progenitor cells may be transplanted or they may be induced to produce healthy differentiated cells for transplant.
In several tissues, stem cells have been isolated and characterised in an attempt to develop new therapies to repair or replace damaged tissues. For example, neural stem cells have been isolated from the mammalian brain (Reynolds and Weiss, Science 255:107 (1992)) and these cells were shown to be multipotential and able to differentiate into neurons, astrocytes and oligodendrocytes. WO 93/01275, WO 94/16718, WO 94/10292 and WO 94/09119 describe uses for these cells.
WO 95/13364 reports the delivery of growth factors to the ventricles of the CNS in order to stimulate neural stem cells to proliferate and produce neural progenitor cells which will develop into neurons, oligodendrocytes or astrocytes. This procedure has many complications which must be addressed before it may be used clinically. Differentiating the target neural stem cells or neural progenitor cells into a desired type of tissue which is functional is one complication. Another complication is choosing a growth factor which does not cause side effects in other areas of the brain.
These publications are limited to isolating or using adult stem cells from the brain (in particular, the tissue around the brain ventricles, the ventricle ependyma, which is the remnant of the embryonic brain germinal zone). Although these publications suggest that progenitor cells may be isolated from the adult peripheral nervous system (“PNS”), the publications define the PNS as the system which originates from the neural crest. There is no reported isolation of a stem cell from the PNS which does not originate from the neural crest.
There are no clinical treatments involving transplants of neural stem cells or neural progenitor cells isolated from the brain nor are there clinical treatments using differentiated cells produced from the neural stem cells or neural progenitor stem cells isolated from the brain. There are also no clinical treatments to endogenously stimulate the neural stem cells or neural progenitor cells of the brain in vivo to produce differentiated cells. Even if there were clinical procedures to transplant fetal neural stem cells or neural progenitor cells from the brain, or to transplant cells differentiated from these stem cells or progenitor cells (e.g. dopaminergic neurons into Parkinson's disease patients), this would not overcome the many problems of transplants from one human to another. As mentioned above, the only current, accessible human source for these neural stem cells and neural progenitor cells is aborted human fetuses, raising serious ethical concerns. Heterologous transplants are also very risky and complicated because of problems with graft rejection, immunosuppression, and the potential for donor grafts transferring diseases or disorders to a recipient. Encapsulation of cells in microspheres has the potential to decrease the likelihood of graft rejection, but this effect is lost if the integrity of the microsphere is disrupted. There is a clear need for safer tissue grafts which can be transplanted to a recipient without being rejected.
The safest type of tissue graft would be one that comes from self (an autologous tissue source). Autologous tissue sources are widely used in procedures such as bone transplants and skin transplants because a source of healthy tissue is readily accessible for transplant to a damaged tissue site. In brain diseases, such as Parkinson's disease, healthy dopaminergic neuronal brain tissue may exist at other sites in the brain but attempts to transplant these neurons would harm the site where the healthy neurons originate. Neural stem cells or neural precursor cells that can be differentiated into dopaminergic neurons may be available at the damaged site or at other sites from which they may be transplanted, but the CNS, particularly the brain, is physically difficult to access. It would be impractical or impossible to access brain or other CNS tissue for biopsy and then again for transplant in patients with weakened health. It would be very useful if there were accessible stem cells or progenitor cells that could be differentiated into CNS cell types, such as dopaminergic neurons, to provide a source of cells for autologous transplants.
It would be useful if neural stem cells or progenitor cells could be identified and isolated outside the CNS and outside the PNS which originates from the neural crest. Medical treatments could then be developed using those neural stem cells, neural progenitor cells or cells differentiated from those cells. It is clear that despite the work that has been done to attempt to treat neurodegenerative diseases by tissue transplant, a need still exists for a pharmaceutical composition in which (1) the composition is accepted by the patient, thus avoiding the difficulties associated with immunosuppression, (2) the composition is safe and effective, thus justifying the cost and effort associated with treatment, (3) the composition provides long term relief of the symptoms associated with the disease, (4) the composition is efficacious during and after transplantation and (5) there are no objections to the ethics of the composition's use.
Thus, there is a clear need to develop neural stem cell cultures or neural progenitor cell cultures from accessible tissues of the PNS which can act as a source of cells that are transplantable to the CNS, PNS, spinal cord or other tissues in vivo in order to replace damaged tissue.